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- PDB-7p8v: The structure of E. coli MutL bound to a 3' resected DNA end -

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Basic information

Entry
Database: PDB / ID: 7p8v
TitleThe structure of E. coli MutL bound to a 3' resected DNA end
Components
  • DNA mismatch repair protein MutL
  • Primer strand
  • Template strandTranscription (biology)
KeywordsDNA BINDING PROTEIN / DNA mismatch repair / Protein-DNA complex
Function / homology
Function and homology information


single-stranded DNA-dependent ATP-dependent DNA helicase complex / mismatch repair involved in maintenance of fidelity involved in DNA-dependent DNA replication / mismatch repair complex / regulation of DNA recombination / nucleotide-excision repair, DNA duplex unwinding / mismatched DNA binding / ATP-dependent DNA damage sensor activity / mismatch repair / ATP hydrolysis activity / DNA binding ...single-stranded DNA-dependent ATP-dependent DNA helicase complex / mismatch repair involved in maintenance of fidelity involved in DNA-dependent DNA replication / mismatch repair complex / regulation of DNA recombination / nucleotide-excision repair, DNA duplex unwinding / mismatched DNA binding / ATP-dependent DNA damage sensor activity / mismatch repair / ATP hydrolysis activity / DNA binding / ATP binding / identical protein binding
Similarity search - Function
DNA mismatch repair protein, MutL / MutL, C-terminal domain, regulatory subdomain / MutL C terminal dimerisation domain / MutL, C-terminal, dimerisation / MutL, C-terminal domain superfamily / MutL, C-terminal domain, dimerisation subdomain / MutL C terminal dimerisation domain / DNA mismatch repair protein family, N-terminal / DNA mismatch repair protein, S5 domain 2-like / DNA mismatch repair, conserved site ...DNA mismatch repair protein, MutL / MutL, C-terminal domain, regulatory subdomain / MutL C terminal dimerisation domain / MutL, C-terminal, dimerisation / MutL, C-terminal domain superfamily / MutL, C-terminal domain, dimerisation subdomain / MutL C terminal dimerisation domain / DNA mismatch repair protein family, N-terminal / DNA mismatch repair protein, S5 domain 2-like / DNA mismatch repair, conserved site / DNA mismatch repair protein MutL/Mlh/Pms / DNA mismatch repair protein, C-terminal domain / DNA mismatch repair proteins mutL / hexB / PMS1 signature. / DNA mismatch repair protein, C-terminal domain / Histidine kinase-, DNA gyrase B-, and HSP90-like ATPase / Histidine kinase/HSP90-like ATPase superfamily / Ribosomal protein S5 domain 2-type fold, subgroup / Ribosomal protein S5 domain 2-type fold
Similarity search - Domain/homology
PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER / DNA / DNA (> 10) / DNA mismatch repair protein MutL
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
DNA molecule (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.6 Å
AuthorsBorsellini, A. / Lamers, M.H.
Funding supportEuropean Union, 1items
OrganizationGrant numberCountry
Marie Sklodowska-Curie Actions, FragNET ITNEuropean Union
CitationJournal: Nucleic Acids Res / Year: 2022
Title: MutL binds to 3' resected DNA ends and blocks DNA polymerase access.
Authors: Alessandro Borsellini / Joyce H G Lebbink / Meindert H Lamers /
Abstract: DNA mismatch repair removes mis-incorporated bases after DNA replication and reduces the error rate a 100-1000-fold. After recognition of a mismatch, a large section of up to a thousand nucleotides ...DNA mismatch repair removes mis-incorporated bases after DNA replication and reduces the error rate a 100-1000-fold. After recognition of a mismatch, a large section of up to a thousand nucleotides is removed from the daughter strand followed by re-synthesis. How these opposite activities are coordinated is poorly understood. Here we show that the Escherichia coli MutL protein binds to the 3' end of the resected strand and blocks access of Pol I and Pol III. The cryo-EM structure of an 85-kDa MutL-DNA complex, determined to 3.7 Å resolution, reveals a unique DNA binding mode that positions MutL at the 3' end of a primer-template, but not at a 5' resected DNA end or a blunt DNA end. Hence, our work reveals a novel role for MutL in the final stages of mismatch repair by preventing premature DNA synthesis during removal of the mismatched strand.
History
DepositionJul 23, 2021Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jun 29, 2022Provider: repository / Type: Initial release
Revision 1.1Aug 31, 2022Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation_author.identifier_ORCID

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: DNA mismatch repair protein MutL
B: DNA mismatch repair protein MutL
C: Template strand
D: Primer strand
hetero molecules


Theoretical massNumber of molelcules
Total (without water)147,8558
Polymers146,7944
Non-polymers1,0614
Water0
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein DNA mismatch repair protein MutL /


Mass: 68005.508 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (strain K12) (bacteria)
Strain: K12 / Gene: mutL, b4170, JW4128 / Production host: Escherichia coli (E. coli) / References: UniProt: P23367
#2: DNA chain Template strand / Transcription (biology)


Mass: 6841.428 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) DNA molecule (others)
#3: DNA chain Primer strand


Mass: 3941.584 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) DNA molecule (others)
#4: Chemical ChemComp-ANP / PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER


Mass: 506.196 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C10H17N6O12P3 / Comment: AMP-PNP, energy-carrying molecule analogue*YM
#5: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg
Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

Component
IDNameTypeEntity IDParent-IDSource
1E. coli MutL N-terminal dimer bound to a 3' resected DNA endCOMPLEX#1-#30RECOMBINANT
23' resected DNA endCOMPLEX#2-#31RECOMBINANT
3E. coli MutL N-terminal dimerCOMPLEX#11RECOMBINANT
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
23Escherichia coli K-12 (bacteria)83333
32DNA molecule (others)2853804
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
23Escherichia coli (E. coli)562
32synthetic construct (others)32630
Buffer solutionpH: 8.5
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMtrisC4H11NO31
2150 mMsodium chlorideNaClSodium chloride1
35 mMmagnesium chlorideMgCl21
42 mMDithiothreitolC4H10O2S21
50.01 %tween20C58H114O261
SpecimenConc.: 0.4 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R0.6/1
VitrificationInstrument: LEICA PLUNGER / Cryogen name: ETHANE / Humidity: 76 % / Chamber temperature: 277.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Calibrated magnification: 105000 X / Nominal defocus max: 2000 nm / Nominal defocus min: 800 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingElectron dose: 54 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 2 / Num. of real images: 539000
EM imaging opticsEnergyfilter slit width: 20 eV

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Processing

SoftwareName: REFMAC / Version: 5.8.0267 / Classification: refinement
EM software
IDNameVersionCategory
2EPUimage acquisition
4Gctf1.06CTF correction
10RELION3.1initial Euler assignment
11RELION3.1final Euler assignment
12RELION3.1classification
13RELION3.13D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 539000
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 149000 / Symmetry type: POINT
RefinementResolution: 3.7→219.9 Å / Cor.coef. Fo:Fc: 0.865
RfactorNum. reflection% reflection
Rwork0.40535 --
obs0.40535 439497 100 %
Displacement parametersBiso mean: 93.808 Å2
Baniso -1Baniso -2Baniso -3
1-4.04 Å2-1.47 Å2-1.4 Å2
2---0.77 Å2-0.64 Å2
3----3.27 Å2

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